Metallothionein mRNA Expression and Cadmium Tolerance in Metal-stressed and Reference Populations of the Springtail <Emphasis Type="Italic">Orchesella cincta</Emphasis>

Metal contamination in soil ecosystems is a permanent and often strong selection pressure. The present study investigates metal tolerance in 17 Orchesella cincta (Collembola) populations from metal-contaminated and reference sites, and combines analyses at the phenotypic and molecular level. Metal tolerance was phenotypically assayed by measuring survival times of laboratory cultures during exposure to cadmium. Comparisons of survival curves showed that five out of eight metal-stressed populations tested evolved increased cadmium tolerance (Stolberg, Plombieres, Hoboken, Hygum and Gusum). In addition, the role of the metallothionein (MT) gene in cadmium tolerance of O. cincta was studied by means of quantitative RT-PCR. The constitutive and Cd-induced MT mRNA expression of the laboratory cultures was measured. Results show that the mean constitutive MT mRNA expression of populations from polluted sites was significantly higher than of populations from reference sites. However, no correlation between MT mRNA expression levels after laboratory exposure to cadmium and field cadmium concentrations was observed. Furthermore, no relation between survival rate during exposure to cadmium and MT mRNA expression was detected. Our results suggest that constitutive MT mRNA expression plays a role in early protection against cadmium toxicity, and indicate that mechanisms other then MT up-regulation are involved in tolerance to prolonged exposure to cadmium.     

Quantitative changes in metallothionein expression in target cell-types in the gills of turbot (Scophthalmus maximus) exposed to Cd, Cu, Zn and after a depuration treatment

Turbot (Scophthalmus maximus) were exposed to two sublethal concentrations (1 and 10 mg metal/l) of cadmium (8.9 and 89 microM Cd), copper (15.26 and 152.6 microM Cu) and zinc (15.3 and 153 microM Zn) for 7 days, and afterwards were maintained depurating for 14 days. Immunoreactive metallothioneins (irMTs) and metal ions were localized in the branchial epithelium by immunohistochemistry (using an anti-Cod MT antibody) and autometallography (AMG), respectively. Metal ions were demonstrated by AMG as black silver deposits (BSD), mainly in mucocytes (MC) and to a lesser extent in the other branchial cell-types (respiratory cells (RC), chloride cells (CC) and basal layer cells (BLC)). Irrespective of the metal supplied, BSD were rapidly visualized in MC after 1 h of exposure. This accumulation did not increase with increasing exposure time and concentration. Metallothionein expression was mainly observed in mature CC in the interlamellar space for all exposure conditions and it was shown that all mature cells express the same amount of irMT. The number of CC exhibiting irMT in metal-exposed turbots increased following short exposure times (1 h-1 day) in the filament epithelium and following longer exposure times (1-7 days) in the secondary lamellae. Total levels of irMT in the gills (quantified by image analysis and densitometry) increased significantly in metal-exposed turbot and were related to increased exposure times. It can be concluded that the total content of irMT in the gills of metal-exposed turbot is governed by changes in the number of mature CC expressing the protein. The quantification of total irMT in branchial CC can be considered as a reliable biomarker of metal exposure since reflects changes in metal bioavailability. This approach based on cell-selective immunohistochemistry can be simplified by only quantifying the number of mature CC. In addition, the dramatic increase of CC in the gills that produces epithelial thickening of the FE enhances migration of CC up to the edge of the SL and provokes the hypertrophy and fusion of secondary lamellae can be considered as unspecific biomarkers of effect indicating disturbed health in turbot.     

PCB77 (3,3',4,4'-tetrachlorobiphenyl) co-exposure prolongs CYP1A induction, and sustains oxidative stress in B(a)P-exposed turbot, Scophthalmus maximus, in a long-term study

Cytochrome P4501A (CYP1A), benzo(a)pyrene (B(a)P) activation and biliary elimination, phase II activities, and peroxisomal and antioxidant activities of turbot (Scophthalmus maximus) were studied in a long-term controlled experiment. Fish were serially exposed in water on day 1 and on completion of months 3, 6 and 9 to 0.1, 0.2, 0.1 and 0.1mg B(a)P/l, respectively, while another group was identically treated with additional PCB77 (3,3',4,4'-tetrachlorobiphenyl) at 1% of concomitant B(a)P (w/w). Temporally persistent responses were obtained by sampling on week 3 and 3 months from each latest exposure. Serial exposure to B(a)P+PCB77 progressively induced liver 7-ethoxyresorufin O-deethylase (EROD) activity and CYP1A protein levels (ELISA, western blotting) towards months 9, 12 and gill EROD activity on month 12. It associated with an apparent increase in liver benzo(a)pyrene diol epoxide (BPDE)-DNA adduct levels (ultrasensitive enzyme radioimmunoassay), and elevated bile B(a)P metabolite levels on month 9 females as compared to males. In contrast, B(a)P alone did not cause (p>0.05) comparable effects on liver EROD, CYP1A, adducts nor on bile metabolites. Both exposed groups demonstrated evidence for lasting oxidative stress as hepatic superoxide dismutase, catalase and glutathione peroxidase activities were significantly altered (p<0.05) with symptomatic pro-oxidant associations among them. Both treatments affected liver somatic index similarly (increase on month 3, decrease on month 9 in males). Continued exposure on month 18 (0.2mg B(a)P/l, 1% PCB77) followed by sampling 6 months later showed sustained induction (p<0.001) of hepatic EROD in B(a)P+PCB77 group, which was not seen in B(a)P alone treatment. Thus, PCB77 co-exposure prolonged CYP1A induction and contributed to a persistent oxidative challenge in B(a)P-exposed turbot. The results indicate synergistic effects of polycyclic aromatic hydrocarbon (PAH) and polychlorinated biphenyl (PCB) exposure in the aquatic environment.     

Pharmacokinetics of a discontinuous absorption process of oxolinic acid in turbot,Scophthalmus maximus,after a single oral administration

1. The pharmacokinetics of oxolinic acid have been studied in 500?g turbot (Scoph thalmus maximus). The fish were kept in seawater at 16 °C with a 15 h/9 h photoperiod. Oxolinic acid was administered orally via a stomach tube at a single dose of 10?mg/kg of body weight. Serum concentrations of oxolinic acid were determined by a (HPLC) using liquid phase extraction with an internal standard and a fluorescence detection. 2. The pharmacokinetic process was not significantly sex-influenced. The short elimination phase of the oxolinic acid in turbot after oral administration was similar to the elimination after intravascular administration. The serum concentration profile of oxolinic acid was better described by a discontinuous absorption model than by compartment models using continuous absorption processes. The absorption of oxolinic acid in turbot was characterized by two distinct phases after a lag time of about 2 h. A time (T_max) of 12 h max was necessary to reach the peak serum concentration (C_max) of 1.41μg/ml. The oral max bioavailability was 27.9%. 3. Based on the minimum inhibitory concentration for susceptible strains, and especially Vibrio anguillarum, the oxolinic acid could be effective in turbot after an oral treatment of 10?mg/kg/day.     

Cellular biomarkers of exposure and biological effect in hepatocytes of turbot (Scophthalmus maximus) exposed to Cd, Cu and Zn and after depuration

Cellular biomarkers of exposure and biological effects were measured in hepatocytes of turbot exposed to either Cd, Cu or Zn at concentrations of 1 and 10 mg/l seawater for 7 days and after depuration for 14 days. Metal content in hepatocyte lysosomes was determined by image analysis after autometallography (AMG) as volume density of autometallographed black silver deposits (Vv(BSD)). Metallothionein (MT) levels were quantified on liver sections by microdensitometry after immunohistochemical staining with a polyclonal anti cod-MT antibody (MT-OD), and in the cytosolic fraction of hepatocytes by difference pulse polarography (MT-DPP). Lysosomal structural changes (lysosomal volume, surface and numerical densities--Vv(LYS), Sv(LYS) and Nv(LYS-), and surface-to-volume ratio S/V(Lys)) were quantified by image analysis after demonstration of beta-glucuronidase activity on liver cryotome sections. Vacuolisation produced by metal-exposure in hepatocytes was quantified by stereology as volume density of vacuoles (Vv(VAC)). Exposure time and metal concentrations significantly affected Vv(BSD) in lysosomes, MT levels and the degree of vacuolisation after 1 h and 1 day exposure to the three metals. The highest Vv(BSD), MT and Vv(VAC) values were recorded after 7 days exposure in all cases. MT-OD and MT-DPP were significantly correlated with Vv(BSD). Vv(LYS) in hepatocytes increased significantly after exposure to the metals. Exposure biomarkers returned to control values after depuration with the exception of those turbots that had been exposed to 10 mg Cd/l. Alike, Vv(LYS) and Sv(lys) (Cu exposure) and Nv(LYS) (Cd and Zn exposures) returned to control values after depuration. It has been therefore demonstrated that the biomarkers used are reversible and return towards control levels once metal exposure ceases. Overall, it is concluded that Vv(BSD), MT-levels and lysosomal responses are valuable biomarkers to assess metal exposure and its effects in turbot, although in quantitative terms the biomarker response varied between metals.     

Environmentally realistic exposure to weathered North Sea oil: Sublethal effects in Atlantic cod (Gadus morhua) and turbot (Scophthalmus maximus)

With increasing oil and gas activities and transport in the Arctic, there is a need to understand how operational or accidental releases of substances affect marine organisms from a pristine environment. The aim of the current study was to describe and compare the responses of two marine fish species, Atlantic cod (Gadus morhua) and turbot (Scophthalmus maximus), following exposure to three levels (low, medium, high) of the water-soluble fraction of a North Sea crude oil for 16 days. The exposure system simulated environmental exposure by allowing clean seawater to percolate through gravel covered in weathered oil before being introduced to aquaria. Both polycyclic aromatic hydrocarbon (PAH) metabolite bile concentrations and cytochrome P4501A (CYP1A) levels and activity increased markedly in comparison with controls in both species, but there were no significant differences between the three exposures. Turbot possessed 4–5-fold higher concentrations of two PAH bile metabolites compared to Atlantic cod by day 8. In contrast, hepatic CYP1A activity in cod was consistently 2–6-fold higher than in turbot with increasing differences over the experimental period. Baseline DNA strand breaks in lymphocytes and kidney cells were low in both species, but was elevated for all treatments by day two. There were no marked indications of the treatments affecting immune functions in either species. This investigation demonstrated that there may be significant differences in responses between species receiving identical exposures and that DNA strand breaks in lymphocytes and kidney cells are sensitive to confinement stress. Data also indicate that some species, such as turbot, may adapt to treatments within days and weeks.     

Comparison of protein expression in plasma from nonylphenol and bisphenol A-exposed Atlantic cod (Gadus morhua) and turbot (Scophthalmus maximus) by use of SELDI-TOF

The overall objective of this study was to compare the expression of plasma proteins in juvenile cod and turbot after a 3 week exposure to two different chemicals known to be estrogenic: 4-nonylphenol (NP, 29 microg/L) and bisphenol A (BPA, 59 microg/L). ProteinChip) array technology in combination with surfaced enhanced laser desorption ionisation-time of flight (SELDI-TOF) was used to investigate general responses in plasma proteins. In addition, an indirect enzyme-linked immunosorbent assay (ELISA) was used to analyse two specific biomarkers of estrogenic exposure, vitellogenin (Vtg) and zona radiata protein (Zrp) in plasma. Both methods revealed clear species specific responses. In cod, 67% of significantly altered proteins showed the same response (up or down regulated) in NP and BPA exposed animals (males and females combined). The rest were either specific to NP (10%), BPA (19%) or they showed opposite responses to the two chemicals (4%). In contrast, only 20% of significantly altered proteins were common for NP and BPA exposed turbot: 60% were altered only in NP and 17% only in BPA. Furthermore, in BPA exposed cod, 77% of the responses were common for male and females, whereas turbot showed only 21% similarity for the two genders. However, NP exposed male and female turbot showed 88% similarity in responses. As gender was not determined in NP exposed cod, gender specific responses could not be determined. ELISA results supported that cod responded clearly to both chemicals as a large increase was observed in Vtg and Zrp levels. Turbot responded strongly to NP, but seemed only slightly affected by BPA. Overall, the results indicated that cod are more sensitive or respond with less specificity to estrogenic chemicals than turbot. The relatively large degree of common responses in NP and BPA exposed cod may indicate that in cod BPA have similar mode of action as NP. Generally, the results show the potential of SELDI-TOF as a tool for comparing multiple responses, and for identifying exposure as well as gender specific responses.     

Effect of Extended Storage Time on the Toxicity of Sediment-Associated Cadmium on Midge Larvae (Chironomus tentans)

The effect of the duration of spiked sediment storage on cadmium toxicity was studied. Sediment samples were spiked with cadmium to obtain concentrations of 0.6,16.0, 29.0 and 53.0 g Cd per g sediment (dry weight). The spiked sediment was then stored in sealed plastic containers at 4°C in the dark. Sediment bioassays, using Chironomus tentans, were conducted immediately and at periodic intervals for up to 4 months. Though the levels of cadmium in the bulk sediment samples from different stored periods were not significantly different, different toxicity levels to C. tentans were observed. The toxicity was significantly different between subsequent storage times. There was a significant decrease in the bioaccumulation factor (BAF) values with extended storage times, indicating a reduction in the bioavailability of cadmium. This study suggests that the storage of spiked sediment used in sediment toxicity study can influence the results.     

The Importance of Metallothionein for the Accumulation of Copper,Zinc and Cadmium in Environmentally Exposed Perch,Perca fluviatilis*

Abstract: A field study of the role of metallothionein (MT) in the binding of heavy metals in perch (Perca fluviatilis), exposed to moderate levels of copper, zinc and cadmium, was performed. Perch were sampled at four sites in Sweden in September during two consecutive years. Two sites were located in the vicinity of a brassworks and two outside the emission range. The first year, fish from the two brassworks sites and from one of the uncontaminated sites were collected. The second year, fish from the most contaminated site and from the two uncontaminated sites were caught. The levels of hepatic copper, zinc and cadmium reflected the concentrations of these metals in water and were increased in fish from the two contaminated sites. The level of cadmium in liver was relatively low. MT was induced in liver of perch caught at the most contaminated site. The hepatic MT content in individual livers correlated well to the accumulation of copper (r=0.85, P<0.001) and zinc (r=0.75, P<0.001). There was a low but significant correlation between the levels of MT and cadmium in the liver (r=0.48, P<0.001). The relationship between MT and metals was very similar both years. Subcellular fractionation of the metals in the liver revealed that an induction of MT was followed by an increased amount of copper, zinc and cadmium bound to the protein. The relative fraction of the total hepatic copper and cadmium bound to MT was increased at the most contaminated site, whereas there was no difference in subcellular distribution of zinc between the sites. In perch from the most contaminated site, the portions of hepatic copper, zinc and cadmium found in the cytosolic fraction were 70, 57 and 81%, respectively. Seventy-one % of the copper, 29% of the zinc and 84% of the cadmium found in hepatic cytosol of fish from the same site, eluted together with MT after gel filtration chromatography. The analysis of the subcellular distribution of copper, zinc and cadmium demonstrates that MT is responsible for the binding of a large amount of the total hepatic cellular content of copper and cadmium in perch.     

Tolerance and pharmacology of ciprostene, a stable epoprostenol (prostacyclin) analogue in humans

Safety, tolerance, and pharmacology of 9-beta-methylcarbacyclin calcium (ciprostene calcium) was investigated in healthy male volunteers. This stable prostacyclin analogue was infused intravenously into groups of 12, 11, and three volunteers for three, six, and eight hours, respectively, in doses up to 480 ng/kg/min. Based on the tolerance data obtained, a single-blind, placebo-controlled study was conducted. Seven subjects were infused for 8 hr/d for three days with ciprostene at a maximum dose of 160 ng/kg/min and seven subjects received placebo. One subject from each group did not complete the infusion schedule, and they were not included in the final analysis. During infusion of ciprostene, consistent changes in blood pressure and heart rate did not occur. Most frequent adverse drug reactions consisted of headache, restlessness, nausea, perspiration, flushing, and jaw pain. As compared with placebo, ADP-induced platelet aggregation was inhibited during the infusion period (P = .048). Significant (P = .04) elevations of platelet cyclic AMP were observed in subjects during infusion of ciprostene. Pre- versus postinfusion routine laboratory evaluations, fibrinogen concentration, antiplasmin activity, and plasminogen and template bleeding times remained unchanged. Placebo- and drug-treated subjects had a daily postinfusion shortening of euglobulin clot lysis time (ECLT). The preinfusion minus postinfusion ECLT for ciprostene subjects on days 2 and 3 (133 and 118 min, respectively) compared with placebo (239 and 217 min) suggest a trend to increased fibrinolytic activity. Based on the outcome of this trial, it is estimated that ciprostene is about 15 times less potent than prostacyclin.(ABSTRACT TRUNCATED AT 250 WORDS)     

Blood Biochemical and Erythrocytic Morpho-pathological Consequences of Naphthalene Intoxication in Indian Teleost,Anabas testudineus (Bloch)

Anabas testudineus (Bloch) was exposed to 0.71 mg/L and 1.42 mg/L (25 and 50% of LC₅₀ value respectively) naphthalene, a polycyclic aromatic hydrocarbon (PAH), for 21 days. Blood biochemical parameters and erythrocytic morphological alterations were assessed to describe the naphthalene toxicity. Biochemical analysis showed a significant increase in glutamic pyruvic transaminase, GPT (576.7 ± 11.79 and 608.9 ± 12.08 U/L, respectively) and alkaline phosphatase, ALP (12.9 ± 0.69 and 13.4 ± 0.64 U/L, respectively) activities under two doses compared with control. Protein and albumin (ALB) content in blood decreased significantly, in comparison with control value in the tune of 22.67 ± 1.04 and 23.97 ± 1.24 g/dl, respectively and 10.7 ± 0.79 and 11.1 ± 0.67 g/dl, respectively. Erythrocytes showed varied symptomatic morphological changes under naphthalene exposure, which included severe denaturation, swelling in cells, appearance of sickle and tear drop cells, and cellular vacuolation. In particularly, the changes were more prominent under higher naphthalene exposure. Following the results, it has been able to establish that GPT, ALP, protein and ALB, and the morphological manifestations of erythrocytes would be good tools of biomarker in monitoring toxicological paradigm, especially to naphthalene exposure in aquatic bodies.     

hPH-20(hSPAM1)mRNA在前列腺腺癌组织中的表达

目的研究人精子膜基因hPH-20(hSPAM1)mRNA在前列腺腺癌组织中的表达。方法采用逆转录聚合酶链反应(RT-PCR)法检测前列腺腺癌(n=35)、正常前列腺(n=5)组织中hPH-20(hSPAM1)mRNA的表达。结果前列腺腺癌中hPH-20(hSPAM1)mRNA的阳性表达率高于正常前列腺组织,B期阳性表达率低于C+D期,高、中分化阳性表达率低于低、未分化,上述差异均有显著性(P<0.05)。前列腺腺癌hPH-20(hSPAM1)mRNA表达水平高于正常前列腺组织(P<0.05)。结论 hPH-20(hSPAM1)mRNA表达可能与前列腺腺癌的发生、发展有关。     

洛沙坦对肾性高血压大鼠心肌AT_(1a)mRNA、AT_(1b)mRNA表达的影响

目的　观察洛沙坦对大鼠肾性高血压的治疗效果及其对大鼠心肌血管紧张素Ⅱ 1型受体亚型AT1a mRNA、AT1b mRNA表达的调控。方法　改进传统的两肾一夹肾性高血压大鼠实验模型制作方法 ,采用定量反转录聚合酶链式反应 (QRT PCR)方法对AT1a mRNA、AT1b mRNA进行定量。结果　①以丝线细钢丝代替银夹可以成功构造大鼠肾血管性高血压模型。②同假手术组大鼠相比 ,模型组大鼠心脏指数明显高于前者 (P <0 0 5 ) ;而治疗组大鼠心脏指数则无明显差异 (P >0 0 5 )。③肾性高血压形成后 ,模型组大鼠心肌AT1amRNA下降 ,而洛沙坦治疗后治疗组AT1b mRNA显著上升 (P <0 0 5 )。结论　洛沙坦有降压作用 ,也能逆转肾血管性高血压引起的心肌肥厚 ,洛沙坦对AT1亚型mR NA的调控差异 ,提示洛沙坦可能对AT1a和AT1b有受体选择性。     

TLR4、MyD88mRNA在胆脂瘤型中耳炎中的表达

目的建立一种实时荧光RT-PCR方法,定量检测中耳胆脂瘤组织中TLR4、MyD88的表达水平。方法选取中耳胆脂瘤组织25份为对照组(其中13份为炎症组,12份为非炎症组);10份正常外耳道皮肤作为正常组。采用实时荧光RT-PCR与Lightcyeler荧光PCR仪定量检测TLR4、MyD88 mRNA的表达水平;采用2-ΔΔCT法处理实时荧光定量RT-PCR数据。结果炎症组TLR4、MyD88mRNA表达水平较正常外耳道组分别上升了4倍和3倍,且差异有统计学意义(P值均<0.05);炎症组TLR4、MyD88mRNA表达水平较非炎症组分别上升了2倍和1倍,且差异有统计学意义(P值均<0.05)。结论中耳胆脂瘤组织中,尤其是炎症反应明显的胆脂瘤组织,TLR4及其下游信号转导分子的表达量发生改变,提示TLR4-MyD88信号传导途径可能参与了胆脂瘤型中耳炎的发病。     

Expression of proenkephalin (PENK) mRNA in inflammatory leukocytes during experimental peritonitis in Swiss mice

Zymosan- or thioglycollate-induced experimental peritoneal inflammation in mice may serve as a convenient model for investigations of involvement of opioid peptides derived from exudatory leukocytes in the inflammatory processes. During peritonitis, the influx of neutrophils and monocytes/macrophages correlated with a sequential appearance of proinflammatory cytokines (IL-1beta and TNFalpha). After both kinds of stimulation, the expression of PENK mRNA was much higher in exudatory peritoneal leukocytes than its basal level in steady state.     

The mRNA Expression of Cyclo-oxygenase-2 (COX-2) and Vascular Endothelial Growth Factor (VEGF) in Human Breast Cancer

Summary

Aims: There is a growing body of evidence that cyclo-oxygenase 2 (COX-2) plays an important role in carcinogenesis and angiogenesis of human tumours. The present study aims to compare COX-2 expression in human breast cancer and adjacent non-cancerous tissue (ANCT), and to identify any correlation between COX-2 and VEGF expression.

Methods: Total cellular RNA was extracted from frozen breast tissue samples according to standard methodology. The mRNA copy numbers for COX-2 and vascular endothelial growth factor 189 (VEGF-189) were determined 40 infiltrating carcinomas and 40 matched ANCT specimens using quantitative RT-PCR and TaqMan methodology.

Results: The COX-2 mRNA copy number per ixg of RNA was two-fold higher in ANCT compared with the cancerous tissue (p = 0.01). Median mRNA copy number was 5.44 x 10⁶ for ANCT and 2.30 x 10⁶for tumour, (ANCT range: 1 x 10⁶to4.12x 10⁷) (tumour range: 1.29 x 10⁵to1.07x 10⁷).

There was a significant correlation between COX-2 and VEGF-189 mRNA copy numbers in the cancer specimens (correlation coefficient = 0.5528, p = 0.0076).

Conclusions: COX-2 mRNA is overexpressed in both human breast cancer and ANCT. We found higher levels in the matched ANCT which suggests that paracrine effects may be important in the role of COX-2 in mammary carcinogenesis. Furthermore, our results indicate that in human breast cancers COX-2 overexpression is linked to VEGF-189 overexpression and therefore tumour angiogenesis.     

Pharmacological Modulation of Platelet-derived Growth Factor (B) mRNA Expression in Alveolar Macrophages and Adherent Monocytes

Summary: The macrophage profibrotic cytokine, Platelet Derived Growth Factor B [PDGF(B)], is thought to play a central role in orchestrating the fibrotic response in the pathogenesis of cryptogenic fibrosing alveolitis. In this study, we have asked if drugs that increase intracellular cAMP and are commonly administered to patients with lung disease have the ability to downregulate PDGF(B) mRNA. Incubation of human alveolar macrophages from healthy smokers in the presence of dibutyryl cAMP prevented the previously reported dexamethasone-induced increase in PDGF(B) mRNA (P<0.05). Similarly, the combination of aminophylline (2.5 mM) and salbutamol (1 μM) prevented the adherence-dependent increase in PDGF(B) mRNA in adherent human peripheral blood monocytes (P<0.05), whilst causing an increase in the mRNA expression of the cAMP-dependent gene c-fos (P=0.059), and an increase in the intracellular concentration of cAMP (P=0.05). Finally, the presence of a lower concentration of aminophylline (0.25 m) in conjunction with salbutamol (1 μM) also prevented the dexamethasone-induced increase in PDGF(B) mRNA in alveolar macrophages from healthy smokers (P<0.05). Stimulation by these drugs was not associated with a change in the abundance of the mRNA of the house-keeping gene, glyceraldehyde-3-phosphate dehydrogenase. We speculate that drugs, which increase intracellular cAMP, may provide a novel therapeutic avenue whereby PDGF(B) expression in patients with cryptogenic fibrosing alveolitis may be reduced.     

膀胱肿瘤多药耐药基因的表达及其临床意义

目的　探讨膀胱肿瘤多药耐药基因 (MDR- 1m RNA)的表达与临床化疗的相关性。方法　采用逆转录多聚酶链反应 (RT- PCR)技术 ,对 2 6例正常膀胱粘膜和 30例膀胱肿瘤 MDR- 1m RNA的表达进行了检测。结果　 2 6例正常膀胱粘膜中 MDR- 1m RNA表达为 15 .4% ,16例初发膀胱癌 MDR- 1m RNA有表达为37.5 % ,14例复发膀胱癌的 MDR- 1m RNA表达为 78.6 % ,并对其进行了临床生物学行为的观察和比较。结论　多药耐药现象在正常膀胱粘膜中有表达 ,在部分初发膀胱癌中有先天性存在 ,RT- PCR方法检测膀胱肿瘤时具有相对敏感、特异、方便、快速和结果可靠等优点     

Combined Effect of Deoxynivalenol (DON) and Porcine Circovirus Type 2 (Pcv2) on Inflammatory Cytokine mRNA Expression

A host’s immune system can be invaded by mycotoxin deoxynivalenol (DON) poisoning and porcine circovirus type 2 (PCV2) infections, which affect the host’s natural immune function. Pro-inflammatory cytokines, IL-1β and IL-6, are important regulators in the process of natural immune response, which participate in inflammatory response and enhance immune-mediated tissue damage. Preliminary studies have shown that DON promotes PCV2 infection by activating the MAPK signaling pathway. Here, we explored whether the mRNA expression of IL-1β and IL-6, induced by the combination of DON and PCV2, would depend on the MAPK signaling pathway. Specific pharmacological antagonists U0126, SP600125 and SB203580, were used to inhibit the activities of ERK, JNK and p38 in the MAPK signaling pathway, respectively. Then, the mRNA expression of IL-1β and IL-6 in PK-15 cells was detected to explore the effect of the MAPK signaling pathway on IL-1β and IL-6 mRNA induced by DON and PCV2. The results showed that PK-15 cells treated with DON or PCV2 induced the mRNA expression of IL-1β and IL-6 in a time- and dose-dependent manner. The combination of DON and PCV2 has an additive effect on inducing the mRNA expression of IL-1β and IL-6. Additionally, both DON and PCV2 could induce the mRNA expression of IL-1β and IL-6 via the ERK and the p38 MAPK signal pathways, while PCV2 could induce it via the JNK signal pathway. Taken together, our results suggest that MAPKs play a contributory role in IL-1β and IL-6 mRNA expression when induced by both DON and PCV2.